Using the mouse skin initiation/promotion model the contributions of the PGE2 receptors, EP1, EP2, EP3 and EP4 in papilloma development were investigated. Indomethacin was used to inhibit epidermal PGE2 production and reduced papilloma formation about 60% in initiated/promoted mice. In TPA/indomethacin treated mice the EP2 agonist (butaprost) restored papilloma formation to TPA alone treatment levels, but EP1, EP3 and EP4 agonists were ineffective. In papillomas, the EP2 agonist increased Ras and PKA activation, as well as c-AMP, p-Src, p-EGFR, p-Erk and p-Akt levels. Treatment of papillomas with EGFR (Ag1478) or Src (PP2) inhibitors demonstrated that p-EGFR was downstream of p-Src. AG1478 also inhibited the activation of Ras, Erk and AKT. EP2s role in papilloma development was confirmed by the observations that EP2 -/- mice showed a 60% reduction in papilloma numbers and reduced activation of Ras, Src, AKT and Erk. Immunoprecipitation of p-Src or EP2 indicated the presence of an EP2-B-arrestin1/2-p-Src complex in papillomas of wild type. B-arrestin1/2 can contribute to EP2 internalization as well as facilitate GPCR independent EP2-mediated signaling. Thus, the data indicate that PGE2 acting via EP2 activates signal transduction pathways that can contribute to mouse skin papilloma development in the initiation/promotion model. The prostaglandin E2 (PGE2) G protein coupled receptor (GPCR), EP2, plays an important role in mouse skin tumor development. Because keratinocyte replication is essential for tumor development, we investigated EP2-activated signaling pathways that contributed to keratinocyte replication. Topical application of the EP2 agonist, butaprost, dose-dependently increased keratinocyte replication and activated EGFR and PKA;and inhibition of EGFR or PKA (AG1478 or H89) decreased butaprost-induced keratinocyte replication about 70%. Because several previous studies indicated that GPCR activation of EGFR involved a &#946;-arrestin/Src complex, the possibility of EP2 acting via this mechanism in keratinocytes was investigated. In the present study, it was observed that butaprost dose and time-debendently increased EGFR and Src activation and induced &#946;-arrestin1/Src complex formation, and that &#946;-arrestin1-deficiency reduced Src and EGFR activation. In agreement with B-arrestin1 leading to Src and subsequently EGFR activation, studies with Src and EGFR inhibitors (PP2 and AG1478) indicated Src to be upstream of p-EGFR. Butaprost also induced the activation of Akt, ERK1/2 and STAT3;and inhibition of EGFR reduced their activation about 60%, indicating these effectors to be activated, in part, by EGFR. In addition to EGFR activation, butaprost increased cAMP levels and PKA activation in mouse skin as measured by p-GSK3&#946;and p-CREB formation. PKA inhibition (H89) decreased butaprost-induced GSK3B, CREB and ERK activation, but did not affect EGFR activation. Thus, the results indicated that EP2 contributed to mouse skin keratinocyte replication by G protein dependent activation of PKA, and G protein independent, &#946;-arrestin1 dependent, activation of EGFR.